Multiplexed and high-throughput neuronal fluorescence imaging with diffusible probes
Abstract
Synapses contain hundreds of distinct proteins whose heterogeneous expression levels are determinants of synaptic plasticity and signal transmission relevant to a range of diseases. Here, we use diffusible nucleic acid imaging probes to profile neuronal synapses using multiplexed confocal and super-resolution microscopy. Confocal imaging is performed using high-affinity locked nucleic acid imaging probes that stably yet reversibly bind to oligonucleotides conjugated to antibodies and peptides. Super-resolution PAINT imaging of the same targets is performed using low-affinity DNA imaging probes to resolve nanometer-scale synaptic protein organization across nine distinct protein targets. Our approach enables the quantitative analysis of thousands of synapses in neuronal culture to identify putative synaptic sub-types and co-localization patterns from one dozen proteins. Application to characterize synaptic reorganization following neuronal activity blockade reveals coordinated upregulation of the post-synaptic proteins PSD-95, SHANK3 and Homer-1b/c, as well as increased correlation between synaptic markers in the active and synaptic vesicle zones.
Document Details
- Document Type
- Pub Defense Publication
- Publication Date
- Sep 26, 2019
- Source ID
- 10.1038/s41467-019-12372-6
Entities
People
- Anthony B. Kulesa
- Demian Park
- Edward Boyden
- Eike-Christian Wamhoff
- Eric Danielson
- Jeffrey R. Cottrell
- Karen Perez De Arce
- Li Li
- Mark Bathe
- Paul Blainey
- Rémi Veneziano
- Simon Gordonov
- Syuan-ming Guo
Organizations
- National Institutes of Health
- National Science Foundation
- United States Army